JP2771915B2 - Simultaneous automatic analysis system for various compounds with absorption in the ultraviolet and visible regions - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a system for automatically analyzing a plurality of compounds absorbing wavelengths in the ultraviolet and visible regions. More specifically, the present invention relates to the simultaneous and automatic analysis of a plurality of compounds absorbing wavelengths in the ultraviolet and visible regions. The present invention relates to an analysis system capable of performing qualitative and quantitative analysis.

[0002]

2. Description of the Related Art As an analysis system for simultaneously and automatically qualifying and quantifying various kinds of compounds, for example, an analysis system combining a gas chromatograph and a workstation, a high-performance liquid chromatograph, a single wavelength ultraviolet-visible detection method, and the like. An analysis system combining a workstation and an analysis system combining a high-performance liquid chromatograph, a dual-wavelength ultraviolet-visible detection method, and a workstation are known.

As an analysis system for qualitatively and quantitatively determining various kinds of compounds with high precision, for example, an analysis system combining a high-performance liquid chromatograph and a multi-wavelength detection method, and a gas chromatograph and a mass spectrometry detection method are used. An analysis system that combines an appropriate analysis system and an ordinary chemical analysis is known.

[0004]

However, all of the conventional analysis systems have the following problems.
That is, in an analysis system combining a gas chromatograph and a workstation, and an analysis system combining a gas chromatograph and a mass spectrometry detection method, when analyzing volatile compounds and thermostable compounds simultaneously and simultaneously, Is effective, but cannot be applied to hardly volatile compounds and thermally unstable compounds.

An analysis system combining a high-performance liquid chromatograph with a single-wavelength ultraviolet-visible detection method and a workstation, and an analysis system combining a high-performance liquid chromatograph with a dual-wavelength ultraviolet-visible detection method and a workstation are disclosed. It is effective when analyzing compounds having absorption at specific wavelengths simultaneously and simultaneously.However, when the compounds to be analyzed are naturally restricted, when analyzing many kinds of compounds having absorption at various wavelengths, There was a problem that it could not be applied. In addition, this analysis system has a problem that it is difficult to perform highly reliable identification because qualitative information is qualitative based on the retained information or the ratio of the retained information to two wavelengths.

In an analysis system in which a high-performance liquid chromatograph and a multi-wavelength detection method are combined, high-precision analysis can be performed because identification is performed based on spectrum information, but ordinary chemical analysis is appropriately combined. There is a problem that it takes a lot of labor and time as in the analysis of the above.

Accordingly, an object of the present invention is to simultaneously and automatically increase a variety of compounds in a short time, even if they are compounds that have absorption in the ultraviolet and visible regions, even if they are hardly volatile compounds and heat-labile compounds. An object of the present invention is to provide a simultaneous automatic analysis system for various compounds having absorption in the ultraviolet and visible regions, which can perform accurate qualitative and quantitative determinations.

[0008]

Means for Solving the Problems The present inventors have studied various analytical methods in order to solve the above-mentioned problems, and as a result, using a high performance liquid chromatograph, have been able to identify various compounds having absorption in the ultraviolet and visible regions. Each separated compound is separated and detected as a spectrum by a photodiode array detector, and a series of information processing is performed on the retention time, peak area, and spectrum data of each separated compound obtained by the separation and detection. It has been found that the above object can be achieved by applying the method.

The present invention has been made on the basis of the above-mentioned findings, and it has been found that various types of compounds having absorption in the ultraviolet and visible regions and an internal standard substance are separated by a column using a high performance liquid chromatograph and then separated. Detect each compound as a spectrum with a multi-wavelength detector, calculate the retention index of each separation compound from the obtained retention time of each separation compound and the retention time of the retention index substance analyzed in advance, and calculate the retention index of these retention indexes. Qualitatively identifying each compound by searching a spectral library from information and spectral information, and then quantifying each qualitative compound by an internal standard method. An object of the present invention is to provide a simultaneous automatic analysis system for various kinds of compounds.

[0010]

According to the present invention, a high-performance liquid chromatograph is used, usually by using a high-performance liquid chromatograph and a workstation integrated with the high-performance liquid chromatograph, to execute a software program incorporated in the work station, thereby making the ultraviolet-visible liquid chromatograph work. After separating many kinds of compounds having absorption in the part and the internal standard substance by a column, each separated separated compound is detected as a spectrum by a detector, for example, a multi-wavelength detector (photodiode array detector), and obtained. The retention index of each separated compound is calculated from the retention time of each separated compound thus obtained and the retention time of the retention indicator substance analyzed in advance, and a database-based spectrum library is searched from the information of these retention indexes and the information of the spectrum. Can be used to qualify each separated compound, followed by each qualitative separation Objects can be quantified by the internal standard method.

[0011]

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS One embodiment of the present invention will be described below with reference to FIGS.

The analysis system used in the embodiment is as follows.
A high-performance liquid chromatograph having a column for separating various types of compounds having absorption in the ultraviolet and visible regions and a photodiode array detector (resolution: 2 nm) for detecting the spectrum of each separated compound separated by the column, and an integral part thereof The analysis system has a workstation in which a software program for executing information processing described below based on information obtained from each separated compound separated by the column is incorporated.

Thus, the compounds having an absorption in the ultraviolet and visible regions to be analyzed in the present embodiment have a wavelength of 220 to 600.
It is a compound having an absorption at nm. Examples of such a compound include preservatives and bactericides such as benzoic acid and salicylic acid, and U compounds such as oxybenzone and ethyl p-aminobenzoate.
V absorbent, sodium dodecylbenzenesulfonate, surfactant such as lauroylmethyltaurine, acetic acid-dl-α
Antioxidants such as tocopherol and butylhydroxytoluene, pharmaceutically active agents such as guaiazulene and diphenhydramine hydrochloride, pigments such as red No. 2 and yellow No. 403, plasticizers such as dimethyl phthalate and triphenyl phosphate, stearic acid and myristin Examples include oils such as acids, and organic acids such as malic acid and succinic acid.

As the internal standard substance used in the present embodiment, a substance having an absorption at 220 nm can be used. As such a substance, for example, propiophenone, butyrophenone, valerophenone and the like are preferably used. .

Further, as the retention index material used in the present embodiment, for example, Al Kano phenone, gallic acid Al <br/> kill esters are preferably used.

As the column used in the present embodiment, a column comprising a stationary phase and a mobile phase is used. As the stationary phase, for example, silica gel in which an octadecyl group and an octyl group are chemically bonded is preferably used, and silica gel in which an octadecyl group is chemically bonded is more preferable. As the mobile phase, a mobile phase in which the mixing ratio of water and the organic solvent is gradually changed is used. As the water, for example, water to which one or more kinds of sodium perchlorate, sodium chloride, phosphoric acid and the like are added is preferably used. Also, as the organic solvent,
For example, acetonitrile, methanol, ethanol and the like are preferably used, and acetonitrile is more preferred.

In order to carry out the present embodiment, first, the retention indicator substance is separated by a column of a high performance liquid chromatograph (step 1), the retention time is obtained, and this is stored in a software program of a workstation. It is written (step 2).

Next, various compounds having an absorption in the ultraviolet and visible regions to be analyzed and an internal standard substance are subjected to high performance liquid chromatography, and the compound and the internal standard substance are separated by the column (step 3). .

From the chromatogram, the retention times of each compound and internal standard substance are written as retention time information according to a software program, and the written retention time and the retention time of the retention index substance analyzed in advance (FIG. 3 and the following table). ), The retention indices of each compound and the internal standard are calculated according to the following formulas and written into the software program as retention index information (step 4).
1), the ratio between the peak area of each separated compound (a variety of compounds having an absorption in the ultraviolet and visible regions) of the chromatogram and the peak area of the internal standard substance is calculated according to the software program, and the ratio is calculated as the peak area ratio by the software program. Each is written (Step 4-2). Further, each separated compound and internal standard substance are sequentially detected as a spectrum by a photodiode array detector, and written on a soft program as spectrum information (Step 4-3). From the detected spectrum and the chromatogram, for example, three-dimensional spectral data shown in FIG. 2 is obtained.

Table 1 (see the chromatogram shown in FIG. 3) ───────────────────────────── n retention indicator substance retention time ( Min) Difference from n-1 N １ 1 Gallic acid 8.782 8.82 4 2 Methyl gallate 11.938 3.156 5 Ethyl gallate 13.906 1.968 6 n-propyl gallate 15.973 2.067 7 5 acetophenone 18.033 2.060 8 6 n-propiophenone 20.843 2.810 9 7 n-butyrophenone 22.899 2.056 10 8 valerophenone 24.716 1.817 11 9 n-hexanophenone 26.363 1.652 12 10 n-heptanophenone 27.857 1.494 13 11 n-octanophenone 29.240 1.437 14 12 n-decanophenone 31.728 2.479 15 13 laurophenone 33.574 1.846 16 14 n-tetradecanophenone 35.458 1.884 17

[Calculation formula for calculating retention index of each separated compound from retention time of retention index substance analyzed in advance] RI _S : retention index of each separated compound RT _S : retention time of each separated compound RT _n : retention time of retention indicator substance

Next, according to the software program, a substance corresponding to each retention index information is searched from a spectrum library created in advance based on the retention index information and the spectrum information obtained as described above,
Further, the degree of coincidence with the spectrum in the spectral library prepared as a database is calculated, and a substance having a high degree of coincidence is selected as a target substance from the spectral library and identified (step 5).

Thereafter, according to a software program for performing quantification by the internal standard method, a substance corresponding to this compound is selected by searching from a calibration curve library prepared in advance based on information of each identified compound. Then, the content of the target compound is calculated from the calibration curve of the selected substance and the peak area ratio calculated in step 4-2, and the quantification of each substance is completed (step 6).

Finally, according to the software program, the retention time, retention index, substance name, spectrum in the spectrum library, , The amount of the substance, and the chromatogram are output by the printer (step 7).

Therefore, according to the present embodiment, for example, when a high-performance liquid chromatograph and a workstation integrated with the high-performance liquid chromatograph are used to execute a soft program to analyze a cosmetic, a household product, a food, and the like, It is possible to simultaneously and automatically perform high-precision qualitative and quantitative quantification of many types of compounds having absorption in the ultraviolet and visible regions, even if they are non-volatile compounds and heat-labile compounds. And time can be saved.

Next, the present invention will be described more specifically with reference to measurement examples. Needless to say, the present invention is not limited to the following measurement examples.

Measurement Example 1 Under the conditions of the following (1) and (2), 24 compounds having absorption in the ultraviolet and visible regions were subjected to separation and qualification by simultaneous automatic analysis of the present invention, and the result of the following (3) was obtained. (1) Separation conditions Separation was performed using the following columns. Column: Inertsil ODS-2 (stainless steel column) 250mm X 2.1mm φ Eluent: A 100mM sodium perchlorate 0.1% phosphoric acid aqueous solution B Acetonitrile Time program Time (min) A (%) B (%) 0.0 100 0 3.0 100 0 30.0 0 100 40.0 0 100 42.0 100 0 50.0 100 0 flow rate; 0.3 ml / min column temperature; 40 ° C. (2) retention index material and the internal standard substance holding indicator substance; al Kano phenone (C ₂ ~C ₁₄₎ gallate Alkyl ester (C ₁ -C ₄ ) internal standard; butyrophenone (3) Results As shown in Table 2 below and FIG. Table 2 shows the chromatogram shown in FIG.

Table 2 Separation and qualification of 24 compounds having absorption in the ultraviolet and visible regionsピ ー ク Peak number Retention time (min) Retention index Compound match １ 1 8.77 399 HCl Pyridoxine 998 2 10.48 454 Resorcinol 999 3 10.77 463 Methyl nicotinate 999 4 11.70 492 Xylene sulfonate 999 5 12.80 544 Yellow No. 403 995 6 14.17 613 Benzyl alcohol 999 7 15.58 681 Phenoxyethanol 999 8 16.30 716 Anisic acid 998 9 17.51 775 Ethyl p - aminobenzoate 999 10 19.49 852 grayed AIA's Ren Acid 998 11 20.28 880 Pas Raokishi acid n- propyl 997 12 21.15 915 Lauroylmethyl Taurine 999 13 22.14 963 hydrochloric acid Kuroruhekishi di emissions 990 14 23.59 1038 Benzophenone 993 15 24.10 1066 Lauroyl di ethanol ami de 999 16 25.23 1131 n-pro Pirufutareto 997 17 26.64 1219 Ben Zetoniumu Kurorai de 999 18 27.99 1309 anthracene 993 19 28.13 1389 hexachlorophene 998 20 30.01 1431 hexadecyl Kuroroben monkey Koniumu 999 21 30.83 1464 Guaia durene 999 22 32.04 1517 Purple 401 No. 999 23 34.77 1663 n-octyl phthalate 990 24 34.98 1675 n-octyl phthalate 999 25 42.94 1721 α-tocopherol 995

Measurement Example 2 Under the conditions of (1) and (2) below, the qualitative analysis of compounds having absorption in the visible and visible part in the systemic detergent A manufactured by A, the dishwashing detergent B manufactured by b, and the cosmetic C manufactured by c. And quantification were performed by the simultaneous automatic analysis system of the present invention, and the following result (3) was obtained. (1) Separation conditions Separation was performed using the following columns. Column: Inertsil ODS-2 (stainless steel column) 250mm X 2.1mm φ Eluent: A 100mM sodium perchlorate 0.1% phosphoric acid aqueous solution B Acetonitrile Time program Time (min) A (%) B (%) 0.0 100 0 3.0 100 0 30.0 0 100 40.0 0 100 42.0 100 0 50.0 100 0 flow rate; 0.3 ml / min column temperature; 40 ° C. (2) retention index material and the internal standard substance holding indicator substance; al Kano phenone (C ₂ ~C ₁₄₎ gallate Alkyl ester (C ₁ -C ₄ ) internal standard substance; butyrophenone (3) Results As shown in Table 3 below and FIGS. Table 3 shows the chromatograms shown in FIGS.

Table 3 Names and amounts of compounds having absorption in the ultraviolet and visible regions contained in products A, B, and C ───────────Sample peak Retention time Retention index Compound match Weight number (min) (ug) ────────────────────── １ 1 24.94 1114 Oxybenzone 990 0.26 A2 26.59 1215 Trichloro Cal bar Niri de 999 0.38 3 30.18 1436 dibutyl hydroxy toluene 995 0.20 ─────────────────────────────────── 1 10.20 445 p- Toluenesulfonic acid 999 1.5 B 2 15.85 694 Benzoic acid 995 0.64 3 24.13 1068 Lauroyl di ethanol ami de 999 2.1 ─────────────────────────────────── 1 15.61 682 Pas Raokishi benzoate 999 0.65 2 18.36 812 Pas Raokishi benzoate 999 0.19 C 3 21.94 953 Oxybenzone 998 4.8 4 31.18 1478 EHD ^* 990 7.7 5 31.85 1507 EMC ^** 999 7.2 ───────────────────────── ────────── * EHD 2-Ethylhexyl-P-methoxy-cinnamate ** EMC 2-Ethylhexyl-P-dimethyl-aminobenzoate

[0031]

According to the simultaneous automatic analysis system of the present invention, as long as the compound has an absorption in the ultraviolet and visible regions, even if it is a non-volatile compound or a heat-labile compound, various compounds can be simultaneously and automatically analyzed. In addition, highly accurate qualitative and quantitative determination can be performed, and labor and time required for analysis can be saved.

[Brief description of the drawings]

FIG. 1 is a flow chart showing the flow of one embodiment of the simultaneous automatic analysis system of various compounds having absorption in the ultraviolet and visible regions of the present invention.

FIG. 2 is a three-dimensional spectrum diagram showing spectra of various types of compounds obtained by one embodiment of the simultaneous automatic analysis system for various types of compounds having absorption in the ultraviolet and visible regions shown in FIG.

FIG. 3 is a chromatogram of 14 kinds of retention indicator substances.

FIG. 4 is a chromatogram of 24 compounds having absorption in the ultraviolet and visible regions obtained by the simultaneous automatic analysis of the present invention.

FIG. 5 is a chromatogram of a compound having an absorption in the ultraviolet and visible regions in Product A obtained by the simultaneous automatic analysis of the present invention.

FIG. 6 is a chromatogram of a compound having an absorption in the ultraviolet and visible regions in product B obtained by the simultaneous automatic analysis of the present invention.

──────────────────────────────────────────────────続 き Continuation of the front page (56) References JP-A-1-242959 (JP, A) JP-A-1-277754 (JP, A) JP-A 1-161123 (JP, A) JP-A-63- 204146 (JP, A) JP-A-49-50993 (JP, A) (58) Fields investigated (Int. Cl. ⁶ , DB name) G01N 30/86 G01N 30/74

Claims (1)

(57) [Claims] 1. A high-performance liquid chromatograph is used to separate various kinds of compounds having absorption in the ultraviolet and visible regions and an internal standard substance by a column, and each separated compound is detected as a spectrum by a multi-wavelength detector. Calculate the retention index of each separated compound from the obtained retention time of each separated compound and the retention time of the retention index substance analyzed in advance, and search the spectrum library from the information of these retention indexes and the information of the spectrum. A system for automatically analyzing various compounds having absorption in the ultraviolet and visible regions, wherein each of the above compounds is qualitatively determined and then the qualitatively determined compounds are quantified by an internal standard method.